PHORBOL ESTER PMA STIMULATES PROTEIN-SYNTHESIS AND INCREASES THE LEVELS OF ACTIVE ELONGATION-FACTORS EF-1-ALPHA AND EF-2 IN AGING HUMAN FIBROBLASTS

Phorbol esters modulate gene expression, reorganize the cytoskeleton and stimulate bulk protein synthesis and the steps of initiation and elongation. We have observed that a phorbol ester PMA stimulates protein synthesis and increases the amounts of active elongation factors, EF-1alpha and EF-2 in cultured human fibroblasts MRC-5 undergoing ageing. Although bulk protein synthesis slows down during ageing, the cellular response to the stimulatory effects of PMA is higher in senescent cells. Similarly, despite the age-related decline in the amounts of active EF-1alpha and EF-2, senescent cells exhibit a higher response to PMA. The results indicate an age-dependent increase of cellular responsiveness to PMA and provide evidence about both the integrity of the translational apparatus and the effectiveness of the signal transduction pathways during cellular ageing. In comparison, the effects of PMA on esters on total protein synthesis by affecting various components of the protein synthetic machinery are especially intriguing. For example, the stimulatory effect of PMA (4-beta-phorbol-12-beta-myristate-13-alpha-acetate) on protein synthesis has been related to the stimulation of the activities of the initiation factors IF-3, IF-4B and IF-4F [6], a valyl-tRNA transferase [7] and the two elongation factors EF-1alpha and EF-2 [8,9]. The regulation of protein synthesis bears an important role in many cellular and physiological processes like cell growth, cell division, differentiation, ageing and transformation [10,11]. Since protein synthesis declines during ageing at both cellular and organismic levels [11], the modulatory action of PMA on the translational apparatus facilitates the use as a tool for investigating the regulatory mechanisms of protein synthesis in relation to ageing. Our previous studies on age-related changes concerning the cellular levels of elongation factors have documented decreased amounts of active elongation factors EF-1alpha and EF-2 in senescent human fibroblasts [12,13]. In our continuing efforts to understand the mechanisms underlying the regulation of protein synthesis during celullar ageing, we have studied the effects of PMA on total protein synthesis and on the amounts of active EF-1alpha and EF-2 of human fibroblasts undergoing ageing in vitro. Our results show that PMA-treatment of both young and senescent fibroblasts stimulates protein synthesis and increases the amounts of active EF-1alpha and EF-2. Although there is a progressive decline in the rate of total protein synthesis during ageing, the extent of PMA-induced stimulation of protein synthesis either remains unaltered or becomes enhanced. Similarly, the PMA-induced increase in the amounts of EF-1alpha and EF-2 is comparable in both young and old cells, although the basal amounts of both enzymes are lower in senescent fibroblasts. These results indicate that the integrity of protein synthetic machinery is maintained along with unaltered signal transduction pathways mediating the action of phorbol esters during ageing of human cells.